Peripheral reduction of FGFR4 with antisense oligonucleotides increases metabolic rate and lowers adiposity in diet-induced obese mice.

Obesity is a primary risk factor for multiple metabolic disorders. Many drugs for the treatment of obesity, which mainly act through CNS as appetite suppressants, have failed during development or been removed from the market due to unacceptable adverse effects. Thus, there are very few efficacious drugs available and remains a great unmet medical need for anti-obesity drugs that increase energy expenditure by acting on peripheral tissues without severe side effects. Here, we report a novel approach involving antisense inhibition of fibroblast growth factor receptor 4 (FGFR4) in peripheral tissues. Treatment of diet-induce obese (DIO) mice with FGFR4 antisense oligonucleotides (ASO) specifically reduced liver FGFR4 expression that not only resulted in decrease in body weight (BW) and adiposity in free-feeding conditions, but also lowered BW and adiposity under caloric restriction. In addition, combination treatment with FGFR4 ASO and rimonabant showed additive reduction in BW and adiposity. FGFR4 ASO treatment increased basal metabolic rate during free-feeding conditions and, more importantly, prevented adaptive decreases of metabolic rate induced by caloric restriction. The treatment increased fatty acid oxidation while decreased lipogenesis in both liver and fat. Mechanistic studies indicated that anti-obesity effect of FGFR4 ASO was mediated at least in part through an induction of plasma FGF15 level resulted from reduction of hepatic FGFR4 expression. The anti-obesity effect was accompanied by improvement in plasma glycemia, whole body insulin sensitivity, plasma lipid levels and liver steatosis. Therefore, FGFR4 could be a potential novel target and antisense reduction of hepatic FGFR4 expression could be an efficacious therapy as an adjunct to diet restriction or to an appetite suppressant for the treatment of obesity and related metabolic disorders

Allele-Specific Inhibition of Rhodopsin With an Antisense Oligonucleotide Slows Photoreceptor Cell DegenerationAllele-Specific Inhibition of Rhodopsin for adRP

PurposeTo preserve photoreceptor cell structure and function in a rodent model of retinitis pigmentosa with P23H rhodopsin by selective inhibition of the mutant rhodopsin allele using a second generation antisense oligonucleotide (ASO).MethodsWild-type mice and rats were treated with ASO by intravitreal (IVT) injection and rhodopsin mRNA and protein expression were measured. Transgenic rats expressing the murine P23H rhodopsin gene (P23H transgenic rat Line 1) were administered either a mouse-specific P23H ASO or a control ASO. The contralateral eye was injected with PBS and used as a comparator control. Electroretinography (ERG) measurements and analyses of the retinal outer nuclear layer were conducted and correlated with rhodopsin mRNA levels.ResultsRhodopsin mRNA and protein expression was reduced after a single ASO injection in wild-type mice with a rhodopsin-specific ASO. Transgenic rat eyes that express a murine P23H rhodopsin gene injected with a murine P23H ASO had a 181 ± 39% better maximum amplitude response (scotopic a-wave) as compared with contralateral PBS-injected eyes; the response in control ASO eyes was not significantly different from comparator contralateral eyes. Morphometric analysis of the outer nuclear layer showed a significantly thicker nuclear layer in eyes injected with murine P23H ASO (18%) versus contralateral PBS-injected eyes.ConclusionsAllele-specific ASO-mediated knockdown of mutant P23H rhodopsin expression slowed the rate of photoreceptor degeneration and preserved the function of photoreceptor cells in eyes of the P23H rhodopsin transgenic rat. Our data indicate that ASO treatment is a potentially effective therapy for the treatment of retinitis pigmentosa

FGF19 increased metabolic rate in vivo and fatty acid oxidation in hepatocytes in vitro.

<p>Welchol-feeding reduced plasma FGF15 levels in DIO mice treated with or without FGFR4 ASO (n = 8–9/group; A). Subcutaneous infusion of recombinant FGF19 to mice at 100 ng/kg/day raised the plasma FGF19 levels similar to the FGF15 levels observed in FGFR4 ASO treated mice, which was diminished 3 days post-infusion (n = 8/group; B). Neither whole body VO₂ nor heart production rate showed difference between two groups before infusion (C). The infusion of FGF19 caused increases in both VO₂ (D) and heat production rate (E) as compared to the pre-infusion baseline values, which were diminished with the termination of infusion. (F) FGF19 also increased fatty acid oxidation rate in <i>in vitro</i> mouse primary hepatocytes when they were treated with vehicle, 1.0 mM AICAR (as a positive control) or 0.5 ng/ml FGF19 (n = 5/group). Data are expressed as mean ± SEM. *<i>P</i><0.05 and **<i>P</i><0.01 vs. saline group.</p

Summary of the design on the pharmacological studies in DIO mice.

<p>Summary of the design on the pharmacological studies in DIO mice.</p

FGFR4 ASO dose-dependently reduced FGFR4 expression, and lowered adiposity and increased whole body metabolic rate.

<p>DIO mice were treated with saline, control (Cont) ASO or FGFR4 ASO #1 at 25, 50, or 75 mg/kg/week for 13 weeks; a group of lean mice treated with saline was used as normal controls. (A) Upper panel, representative Western blots of liver FGFR4 protein and SRB1 protein (as loading control); lower panel, the quantitative values of liver FGFR4 protein after normalizing to SRB1. (B) Cumulative food intake during the treatment. (C) BW changes during the treatment. (D) Total body fat content at different time points of the treatment. (E) Fat depot weights at the end of the treatment. (F) Plasma β-hydroxybutyrate (3HB) levels at week 0 and week 12. (G) Whole body O₂ consumption rate (VO₂). Data are expressed as mean ± SEM (n = 6–9/group). *<i>P</i><0.05 and **<i>P</i><0.01 vs. saline controls, or as indicated in the figure; ^#<i>P</i><0.05 and ^##<i>P</i><0.01 vs. Cont ASO; NS, not significant.</p

Combination treatment with FGFR4 ASO and rimonabant showed additive reduction of BW and adiposity.

<p>DIO mice were treated with saline, control ASO or FGFR4 ASO #1 at 50 mg/kg/week for 8 weeks. Two and half weeks after the initiation of the treatment, the mice were given daily oral administration of rimonabant (R) at 5 mg/kg or vehicle (V) for 5 weeks. (A) FGFR4 mRNA levels in liver and white adipose tissue (WAT) after 8 weeks of the treatment. (B) Cumulative food intake during the treatment. (C) BW changes during the treatment. (D) Total body fat content at different time points of the treatment. (E) Fat depot weights at the end of the treatment. Data are expressed as mean ± SEM (n = 9/group). *<i>P</i><0.05; **<i>P</i><0.01.</p

FGFR4 ASO treatment induced Cyp7α1 expression-bile acid synthesis-FGF15 expression pathway.

<p>(A) Liver Cyp7α1 protein expression after 4 weeks of treatment with saline, control ASO or FGFR4 ASO #1. Upper panel, representative Western blots of liver Cyp7α1 protein and a non-specific protein band (as loading control); lower panel, the quantitative values of liver Cyp7α1 protein after normalizing to the non-specific protein band. (B) Bile acid pool size, (C) plasma total bile acid levels and (D) ileum FGF15 gene expression after 4 weeks of treatment with saline, control ASO or FGFR4 ASO #1 (n = 5-9/group). (E) Plasma FGF15 levels after 2 and 4 weeks of treatment (n = 9/group). Data are expressed as mean ± SEM. *<i>P</i><0.05 and **<i>P</i><0.01 vs. saline controls; ^##<i>P</i><0.01 vs. Cont ASO.</p